Nowadays, the method of gas chromatography (GC) is carried out at constant temperatures in the form of isothermal GC or with an increase in temperature during the passage of the substances to be separated through the chromatography column in the form of temperature-programmed GC. In GC laboratory systems, the chromatography column is heated uniformly in an oven chamber with temperature gradients as small as possible (air bath oven). The air bath enables the column to be heated rapidly, with temperature increases of up to 100° C./min being possible in commercial systems.
GC is based on the partitioning equilibrium between the mobile phase, i.e. the carrier gas, and the stationary phase, which is in the form of a thin film on the capillary wall in the case of capillary columns. The rate of transport of substances in the column depends only slightly on speed, i.e. the flow of the mobile phase. It is, in particular, the temperature-dependent phase equilibrium of the substances between the stationary and the mobile phase which determines the rate of transport. In isothermal GC, substances are separated only in a narrow phase equilibrium range. The signals from slowly transported substances at excessively low temperatures are very wide due to the long transport times and the diffusion which occurs in the process. Some substances are not transported and remain at the inlet, head, front part of the separation column. Temperature-programmed GC (TPGC) is carried out in such a way that a temperature level at which transport through the column is achieved is established for all the substances.
Given appropriate matching of the carrier gas speed and of the heating rate of the separation column, good separation is achieved over a wide phase equilibrium range. One disadvantage of this method is that the substances are still being transported in the column during heating and are therefore exposed to higher temperatures than those required for substance separation and transport. This effect is particularly relevant in the case of rapid GC separation processes, in which high heating rates are employed. Raising the temperature too quickly leads to a simultaneous reduction in separation efficiency since the substances are exposed to temperatures favorable for separation only within small time windows and hence within short column sections. After this, it is only transportation that occurs in the remaining section of the separation column since the temperatures are then too high for separation processes.
Gas chromatography with a temperature gradient (TGGC) along the separation column is based on an idea from the Russian scientist Zhukhovitskii. If each substance has a temperature that is characteristic thereof, above which transport takes place at a significant speed (often referred to as the running temperature), a gradient from the inlet (high temperature) to the outlet (low temperature) as the mixture of substances flows in has the effect that each of the substances accumulates at the temperature (and hence location) at which said temperature once again falls below the running temperature. In the first phase of TGGC, the separation column acts as a collecting or enrichment system. If the temperature level is then raised with the gradient being maintained, each substance migrates spatially to the outlet since the running temperature shifts progressively in this direction. If the temperature at the outlet is precisely equal to the running temperature, the substance elutes from the column and is passed to the detector.
The difference with respect to TPGC is that each substance is only exposed precisely to the temperature corresponding to its running temperature and is not merely transported onward into high temperature zones. The temperature at which a substance elutes at the outlet of the separation column is therefore systematically lower in TGGC than in TPGC.
Moreover, a central effect and advantage of TGGC is the focusing effect. Since there is a temperature gradient around each substance, substance fractions which have moved somewhat ahead of the main zone are held back by the lower temperature level prevailing there. However, the fractions which are further back are transported more quickly by the somewhat higher temperature. The effect of extended diffusion (longitudinal diffusion) during transportation in the capillary is thus compensated. By virtue of the narrowness of the substance signals, their height is increased, and hence measurement sensitivity and the signal/noise ratio are improved.
Despite the theoretical advantages of TGGC, the concept has not found broader commercial application. Originally, Zhukhovitskii's idea was implemented in short packed separation columns, around which was arranged a mobile oven segment that was moved mechanically from the inlet to the outlet of the separation columns and produced the gradient within the oven section. In some cases, the separation column or separation capillary was of circular design and the oven was moved around in a circle. The original temperature gradient method with a moving oven on packed columns is also referred to as chromathermography. The emerging process of capillary gas chromatography using thin fused silica separation columns or fused silica separation capillaries had proven highly efficient, even in the case of isothermal and, especially, temperature-programmed applications. The central focus of technical development was to optimize air bath gas chromatographs in respect of heating rates and uniformity of temperature. From a technical viewpoint too, transferring the concept of chromathermography from short rigid packed columns to thin and flexible separation capillaries with a length of many meters had to be regarded as difficult to implement. A number of solutions are known in the prior art for managing the problems associated with chromathermographic methods.
Thus, U.S. Pat. No. 3,146,616 describes how, in the chromathermographic method, an electric heating arrangement which supplies the separation column with the respectively required heating power in individual turns of a heating coil is switched progressively in space, replacing a mechanically moved oven.
DE 21 495 08 discloses a simple concentric arrangement of a heating arrangement around the separation column, through which there is a countercurrent flow of a cold fluid, which heats up along its path and thus produces a temperature gradient in the separation column. To release the collected substances, a hot fluid flowing in a co-current is passed into the concentric chamber.
A mechanically complex arrangement for producing a temperature gradient along a 2.2 m long capillary column is furthermore described in U.S. Pat. No. 5,028,243. The column is introduced as a planar structure in the form of a spiral into a fluid channel and its temperature is controlled by a corresponding planar structure comprising a fluid channel and connecting openings and a heating wire extending there. With this arrangement, even very low temperatures (−100° C. is mentioned) can act on the column. Moreover, this publication discloses an arrangement in which a spirally wound heating wire is arranged in a tubular sheath, through the center of which the separation column extends. In addition, a fluid can be passed through the arrangement, e.g. a very cold gas. The desired temperature gradient can be produced by means of a second heating coil with a decreasing coil spacing.
A TGGC apparatus with double-concentric sheathing of the separation capillary is furthermore described in U.S. Pat. No. 5,215,556. A fluid for heat exchange is passed in a co-current relative to the direction of the carrier gas through a first sheath, and a second fluid is passed through the outer sheath in a countercurrent. As a result, a linear temperature gradient is obtained. In this process, the temperature of the separation column or separation capillary is heated directly by the first fluid.
U.S. Pat. No. 5,929,321 describes a chromathermographic arrangement comprising a moving oven. The oven is guided in a precise manner over the separation column and produces the desired local gradients there. The particular aim of the invention is to improve selectivity in conventional gas chromatography processes in the form of a pre-separation.
A double-concentric arrangement comprising a coiled separation column on a holder in a tube is disclosed in U.S. Pat. No. 7,914,612 B2. The arrangement is supposed to be about 10 cm long and encloses a 1 to 5 m long separation column. Once installed in an oven, cold fluid is additionally supplied to produce a temperature gradient.
US 2012/0085148 (A1) discloses an additional system for a conventional gas chromatograph, comprising a looped metal capillary, in which a short conventional fused silica separation column is inserted. The aim of the system development is temperature-programmed gas chromatography with very quick heating and cooling cycles. The application relates to a resistance heater, wherein the gas chromatograph is operated with a resistance heater but without the use of a temperature gradient.
U.S. Pat. No. 5,114,439 likewise describes a coiled arrangement of a resistance-heated capillary column, particularly for mobile uses. The temperature is measured by measuring the resistance, although heating of the separation capillary takes place without a gradient.
In U.S. Pat. No. 5,135,549, four techniques for producing a temperature gradient are presented. There, the use of gradients is generally described in certain configurations, wherein the techniques mentioned describe resistance heating via a coating, in particular a wound heater with a variable winding density of a heating wire, a longitudinally directed coolant flow along a heated capillary with continuous warming up of the coolant and a separation column heated separately to different temperatures.
U.S. Pat. No. 5,808,178 discloses a “flash GC”, wherein a resistance-heated metal sheath capillary, in which the GC column is guided. A cooling trap, through which there is an alternating direction of flow by means of a valve arrangement, is additionally described in this patent. In particular, the problematic influence of temperature differences between the lower and upper capillary turns is mentioned in the description of the patent.
As can be seen from the prior art, implementing a uniform temperature gradient along a capillary column of several meters length is a difficult technical challenge. In particular, the temperature must varied very uniformly, with even short deviations leading to delays in substance transport (if undershot) and hence to distorted signal shapes.
To solve the technical problem, separation capillaries have admittedly also been coated with conductive coatings of decreasing thickness in order to allow differences in temperature adjustment by way of the gradual change in resistance, or the temperature gradient has been produced directly around a metal separation column or separation capillary using a heating wire coil with a continuous increase in winding density. Attempts have also been made to work with a resistance-heated separation column which is sheathed concentrically by a guide tube and in which cold nitrogen is passed in a countercurrent with respect to the carrier gas direction in the guide tube and the heated separation column is cooled more intensely with the still-cold fluid at the outlet than with the already heated fluid at the outlet (cf. PHILLIPS, J. B.; JAIN, V. (1995): On-column temperature programming in gas-chromatography using temperature-gradients along the capillary column. In: JOURNAL OF CHROMATOGRAPHIC SCIENCE 33 (10), pages 541-550; COUDERT, M.; VERGNAUD, J. M. (1971): Retention in gas chromatography obtained with a longitudinal temperature gradient with a constant growth rate. In: JOURNAL OF CHROMATOGRAPHY A 54(1), pages 1-8. DOI: 10.1016/S0021-9673(01)80238-7; Contreras, Jesse A.; Rockwood, Alan L.; Tolley, H. Dennis; Lee, Milton L. (2013): Peak sweeping and gating using thermal gradient gas chromatography. In: JOURNAL OF CHROMATOGRAPHY A 1278, pages 160-165).
Common to all technical solutions hitherto is a high outlay on production. These implementations are not suitable for commercial use. Thus, for TGGC analyses in accordance with the prior art, the separation columns have to be modified manually or mounted laboriously on supports to enable the temperature thereof to be spatially controlled by means of temperature control fluids.
It is therefore the object of the invention to provide a TGGC in which efficient separation can be achieved with commercially available separation capillaries that can be interchanged easily and do not require any special temperature control fluids but allow dynamic temperature control with a gradient and entail the use of only small amounts of energy for temperature control.